Metal centers are essential for enzyme catalysis, stabilizing the active site, facilitating electron transfer, and maintaining the structure through coordination with amino acids. In this study, K238H-AeL nanopores with histidine sites were designed as single-molecule reactors for the measurement of single-molecule coordination reactions. The coordination mechanism of Au(III) with histidine and glutamate in biological nanopore confined space was explored. Specifically, Au(III) interacts with the nitrogen (N) atom in the histidine imidazole ring of the K238H-AeL nanopore and the oxygen (O) atom in glutamate to form a stable K238H-Au-Cl